Protein structure and protein assemblies play a crucial role in disease etiology because protein function is intimately linked to its structure. For example, the common neurodegenerative disorders Parkinson's and Alzheimer's disease result from the misfolding and aggregation of specific proteins. The proteins associated with these diseases resemble prion-like pathogens because they form self-propagating protein assemblies through a dynamic process: individual proteins combine to form small protein assemblies and small protein assemblies combine to form large protein assemblies. This dynamic process involves unfolding and/or rearranging protein and/or protein assembly structures. More generally, many of the fastest growing major diseases in the United States, such as Alzheimer's disease, Parkinson's disease, diabetes mellitus type 2, atherosclerosis, and cancer, involve transiently populated conformations of proteins and their assemblies.
Nuclear magnetic resonance (NMR) and x-ray spectroscopy provide ensemble-averaged structures of proteins and have been invaluable for pharmaceutical research. However, these techniques are not well-suited for elucidating detailed structures of dynamic proteins and their complexes. The existing techniques do not capture co-existing, transiently populated protein conformations because they only measure ensemble-averaged structures of proteins. The inability to elucidate co-existing, transient protein conformations has hampered efforts to develop pharmacological strategies for treating or preventing many diseases because a clear molecular target for drug development cannot be discerned.
Ion mobility spectrometry-mass spectrometry (IMS-MS) has been used for structural characterization of generally small organic and inorganic molecules. Recent advancements in the field have led to equipment modifications that allow IMS-MS to be used for research involving large, macromolecular organic and biological compounds. IMS-MS is capable of revealing transient protein structures, and is well suited to study co-existing, transient conformations of proteins and their complexes because it physically separates analytes that differ in mass and shape within milliseconds. IMS-MS can be accomplished using minute amounts of sample within seconds due to the high sensitivity and speed of MS analysis. However, IMS-MS only measures an orientation-averaged cross section of a protein or a protein assembly. Structural details are not revealed by IMS-MS data, and extracting detailed molecular structures from IMS-MS data alone is challenging.
There are no known methods for extracting de novo detailed molecular structures from measured cross sections alone. Current state of the art methods couple structures from traditional techniques (e.g. NMR) with IMS-MS data analysis. However, this approach cannot exploit the full potential of IMS-MS, which is to elucidate structures for exactly those systems where traditional methods fail. Other methods report structures when computed cross sections for theoretical (average) model structures match experimental data or use experimental cross sections as a “filter” to select a specific structure from a pool of computed model structures; however, these methods suffer from many shortcomings. First, different protein structures can have identical cross sections. Second, the experimental cross section may be the average of the cross sections of distinct structures that interconvert in the experiment. Third, protein dynamics in the IMS-MS experiment depend on the charge state. Therefore, there is a need for improved systems and methods that can extract detailed structural information of molecules from IMS-MS data.